The identification of the molecular basis of melanoma progression and chemoresistance has been hampered by a notorious inter- and intra-tumor heterogeneity, and by the fact that during their transformation into metastatic cells, human melanocytes acquire a large number of genetic and epigenetic alterations in multiple survival and cell death signaling cascades. Understanding which alterations are causative of the disease (and which a byproduct), what are their specific impact at each step of melanoma development (i.e. from pre-malignant to in situ and metastatic lesions), and specifically, how do they affect treatment response, remain major challenges in cutaneous oncology. This proposal will test the hypothesis that inactivation of the mitochondrial cell death pathway provides a critical survival advantage during human melanocyte transformation, invasion and metastasis, and that this inactivation has to be compensated or bypassed to overcome melanoma chemoresistance. To account for tumor heterogeneity, we will take advantage of a large melanoma tumor bank generated at the University of Michigan to perform a comprehensive analysis of critical mitochondrial apoptotic modulators at all stages of melanoma progression (Aim 1). Special emphasis will be dedicated to pre-malignant and early melanomas, since initiating events in melanoma pathogenesis are largely unknown. In parallel, we will dissect the functional contribution of the mitochondrial pathway to transformation (Aim 2) and tumorigenicity (Aim 3) of human melanocytes and melanoma cells, and to their response to chemotherapeutic agents (Aim 4). In this context, an important consideration of our research design is that melanomas do not arise simply as a consequence of mutations in normal melanocytes, but stromal compartments are important determinants as well. We will therefore engineer melanocytic lesions with specific defects in apoptotic pathways and determine their impact on tumor growth in settings that recapitulate the architecture of human melanoma in vivo. Finally, we will exploit our preliminary studies with pharmacological strategies able to bypass mitochondrial cell death defects, with the ultimate goal of identifying new treatments able to overcome melanoma chemoresistance. Defects in apoptosis are not limited to the control of melanoma as increased cell survival is a general concept in tumor biology. Therefore, the approaches and results of our studies may be relevant to a variety of other aggressive tumor types and may provide the basis for rational design of improved therapies.